Natural genetic control systems are defined as mechanisms that are able to modify the expression of genes depending on the presence or absence of certain specific molecules in the cellular environment.
The expression of genes may be controlled during the phases of transcription, growth and translation of nucleic acids (FIG. 1).
Negative and positive control of transcription are distinguished. In the former case, a protein receptor, called a repressor, prevents the gene from being expressed by attaching to its operator and also prevents the RNA polymerase from starting the transcription on the promoter. This system is very common in bacteria (tetR, varR, lacI). In the latter case, an activator or transcription factor which attaches to the DNA is required for starting the transcription. This system is common in eukaryotes (e.g. hormonal receptors) but it is also found in bacteria, as is the case with the activator ampR that controls the transcription of β lactamase ampC in Gram-negative bacteria such as Citrobacter freundei (Jacobs et al., Cell, Vol. 88 (1997), pp. 823-832) or purR (Schumacher et al., Cell, Vol. 83 (1995) , pp. 147-155). In a similar fashion, the translation may also be controlled by the attachment of a repressor to the mRNA which prevents the ribosome from starting the translation into proteins. In eukaryotes, the growth of mRNA may additionally be controlled during the modification, splicing, transfer or stable phases.
Regulators are protein units that may be in 2 states: either in an active state (ATR) in which they may, by attaching to DNA or mRNA, control the expression of genes in a positive (+) or negative (−) manner and thus control the synthesis of proteins, or in an inactive state (ITR) in which they can no longer attach to a particular nucleotide sequence. Activation is normally due to allosteric transition by a repressor as a result of the presence or absence of a binding agent with a strong affinity at a specific site on the regulator. This is the case with bacterial regulators belonging to the lacI family (Weickert and Adhya, J. Biol. Chem., Vol. 267 (1992) pp. 15869-74) or with hormonal receptors in eukaryotes. Other biochemical phenomena such as oxidation, phosphorylation or glycosylation may also throw light on these regulators.
In this case where the mechanism is attached by a binding agent, regulators have two attachment sites:                one is an attachment site on the DNA, often characterised by an exposed helix β turn β helix α (HTH) motif, characteristic of proteins linked to DNA;        the other is an attachment site on the binding agent.        
The presence of the binding agent, called an effector, on the regulator normally induces a modification of the shape of the latter so as to permit (or on the contrary to prevent) the attachment of this regulator to a specific nucleotide sequence. These effectors thus work as inducers or co-repressors. These most common configurations are shown in FIGS. 2A and 2B.
This is the case with purine, a co-repressor required for the attachment of the dimeric repressor purR which belongs to the lacI family of bacterial transcriptional regulators, to the purF operator (Schumacher et al., Cell, Vol. 83 (1995), pp. 147-155; Schumacher et al., Science, Vol. 266 (1994), pp. 763-770) or with L-tryptophan, a co-repressor of the trp dimer (Otwinowski et al., Nature, Vol. 335 (1988), pp. 321-329).
The majority of the other bacterial regulators of the lacI family, with the exception of purR, suppress a catabolic, non-biosynthetic reaction, the affinity of the regulators for the operator being greater in the absence of the binding agent on the repressor. Thus tetR and varR are dimers that also belong to the lacI family, which intervene in the mechanisms of bacterial resistance to tetracycline and virginiamycin antibiotics respectively. These repressors, which moreover are highly homologous in their sequences, may attach to the operators “tet” and “var” only in the absence of their respective binding agents, tetracycline or derivatives and virginiamycin S considered as inducing effectors (Hillen et al., Ann. Rev. Microbiol., Vol. 48 (1994), p. 345; Namwat et al., J.Bac., Vol. 183 (March 2001), pp. 2025-2031).
In the presence of an attached inducer, the regulators “release” the operator and the suppression is immediately removed. One may observe the synthesis of tetA and varS, transfer molecules in the membrane that catalyse the transfer of antibiotics out of the cell. This system requires very fine control since it controls the toxicity of the transfer molecule which will at the same time prevent the antibiotic from reaching its target (the ribosome) but it may also release other non-specific ions outside the cell, which is also lethal to the cell. TetR must therefore strongly suppress tetA until a low level of tetracycline (Tc) is present.
As for tetR, its three-dimensional structure is known in the presence of Tc as well as in the presence of the operator (Hinrichs et al., Science, Vol. 264 (1994), pp. 418-420, Kisker et al., J. Mol. Biol., Vol. 247 (1995), pp. 260-280; Orth et al., Nature Structural Biol., Vol. 7, (2000), pp. 215-219). This allows the better understanding of the molecular machinery. A precise orientation of a lateral chain of the terminal amino acids is required so as to allow high-affinity contact between the operator segment of the DNA and the regulator. When the tetracycline is attached, the distance centre-to-centre between the two HTH motifs of the dimer of tetR is increased by 5 and this slight structural modification is sufficient to break the high-affinity contact with the operator. Studies in vitro show that the affinity of tetR for the operator in the absence of Tc is 1012 to 1013 M−1 and it falls by a factor of −109 in the presence of Tc. TetR forms the most efficient inducible transcription control system known to date (Orth et al., Nature Structural Biol., Vol. 7 (2000), pp. 215-219).
In addition, it should be noted with regard to homology of sequence, six classes (A, B, C, D, E and G) of tetR are known among Gram-negative bacteria. The genes that code for these receptors are plasmids or they are transferred by transposons and they can all be induced at nanomolar concentrations of tetracycline. The proteins share 29% of identical amino acids between the different classes, which suggests similar 3D structures (Hillen et al., Annu. Rev. Microbiol., Vol. 48 (1994), pp. 345-332; Klock et al., J. Bac., Vol 161 (1985), pp. 326-332).
As for varR, the affinity parameters in the presence and absence of the inducer are not yet known but it is probable that they are of a similar scale. The affinity of the antibiotic for its target, the 50S ribosome, being similar, a large difference between suppression and induction of the membrane transfer molecule is also vital to the cell.
Other less vital repressors such as lacI show a loss of activity of only 105 in the presence of the inducer (Matthews et al., Nature Structural Biol., Vol. 7 (2000), pp. 184-187).
Here again one should note that there are exceptions since ampR attached to the operator may activate transcription in vitro in the absence of a binding agent or in the presence of the binding agent anhydro-Mur Nac tripeptide. On the other hand, in the presence of another binding agent of ampR, UDP-Mur-Nac pentapeptide, transcription is inhibited (Jacobs et al., Cell, Vol. 88 (1997), pp. 823-832). AraC attaches to the DNA in the presence and the absence of the binding agent (L-arabinose) but transcription is only started in the presence of the binding agent following a DNA “loop” (Soisson et al., Science, Vol. 276 (1997), pp. 421-425).
Still other repressors such as MetJ attach to the operator only in the presence of the positively-charged co-repressor (s-adenosylmethionine) which, rather than a change of configuration, induces an electrostatic effect in the regulator, which increases its affinity for DNA by more than 1,000 times (Phillips and Phillips, Structure, Vol. 2 (1994), pp. 309-316).